Hiroshi Hisano, Hiroaki Sakai, Mika Hamaoka, Hiromi Munemori, Fumitaka Abe, Brigid Meints, Kazuhiro Sato, Patrick M Hayes
{"title":"Rapid development of naked malting barley germplasm through targeted mutagenesis.","authors":"Hiroshi Hisano, Hiroaki Sakai, Mika Hamaoka, Hiromi Munemori, Fumitaka Abe, Brigid Meints, Kazuhiro Sato, Patrick M Hayes","doi":"10.1007/s11032-025-01553-5","DOIUrl":null,"url":null,"abstract":"<p><p>Covered barley (<i>Hordeum vulgare</i>) has historically been preferred for malting, as the husk in this plant protects the embryo during harvest and acts as a filter during brewing. Naked barley, which is typically used as food, has the potential to be used in brewing due to recent technical advances, but the grains contain higher levels of β-glucan and polyphenols, which are undesirable in brewing. Introducing the naked trait into brewing cultivars through crossing is time-consuming due to the need to eliminate these undesirable traits. In this study, we rapidly developed naked barley that is potentially suitable for malting by introducing targeted mutations into <i>Nudum</i> (<i>NUD</i>) using CRISPR/Cas9-mediated targeted mutagenesis. The doubled haploid line 'DH120366', which was used as the parental line, was derived from a cross between two covered malting barley cultivars. We generated CRISPR/Cas9-mediated targeted mutagenized barley harboring mutations in <i>NUD</i> via <i>Agrobacterium tumefaciens</i>-mediated transformation and confirmed the presence of mosaic mutations in one individual from among 16 T<sub>0</sub> transformants. We sowed T<sub>1</sub> grains exhibiting the naked trait and sequenced the <i>NUD</i> gene in these T<sub>1</sub> seedlings, identifying two types of mutations. Shotgun high-throughput whole-genome sequencing confirmed the absence of the transgene in at least one <i>nud</i> mutant line following <i>k</i>-mer-based analysis. Cultivation in a closed growth chamber revealed no significant differences in agronomic traits between the <i>nud</i> mutants and the wild type. This study demonstrates the feasibility of rapidly developing naked barley with potential use for malting and brewing by targeting only <i>NUD</i> via targeted mutagenesis.</p><p><strong>Supplementary information: </strong>The online version contains supplementary material available at 10.1007/s11032-025-01553-5.</p>","PeriodicalId":18769,"journal":{"name":"Molecular Breeding","volume":"45 3","pages":"32"},"PeriodicalIF":2.6000,"publicationDate":"2025-03-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11889295/pdf/","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Molecular Breeding","FirstCategoryId":"97","ListUrlMain":"https://doi.org/10.1007/s11032-025-01553-5","RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2025/3/1 0:00:00","PubModel":"eCollection","JCR":"Q1","JCRName":"AGRONOMY","Score":null,"Total":0}
引用次数: 0
Abstract
Covered barley (Hordeum vulgare) has historically been preferred for malting, as the husk in this plant protects the embryo during harvest and acts as a filter during brewing. Naked barley, which is typically used as food, has the potential to be used in brewing due to recent technical advances, but the grains contain higher levels of β-glucan and polyphenols, which are undesirable in brewing. Introducing the naked trait into brewing cultivars through crossing is time-consuming due to the need to eliminate these undesirable traits. In this study, we rapidly developed naked barley that is potentially suitable for malting by introducing targeted mutations into Nudum (NUD) using CRISPR/Cas9-mediated targeted mutagenesis. The doubled haploid line 'DH120366', which was used as the parental line, was derived from a cross between two covered malting barley cultivars. We generated CRISPR/Cas9-mediated targeted mutagenized barley harboring mutations in NUD via Agrobacterium tumefaciens-mediated transformation and confirmed the presence of mosaic mutations in one individual from among 16 T0 transformants. We sowed T1 grains exhibiting the naked trait and sequenced the NUD gene in these T1 seedlings, identifying two types of mutations. Shotgun high-throughput whole-genome sequencing confirmed the absence of the transgene in at least one nud mutant line following k-mer-based analysis. Cultivation in a closed growth chamber revealed no significant differences in agronomic traits between the nud mutants and the wild type. This study demonstrates the feasibility of rapidly developing naked barley with potential use for malting and brewing by targeting only NUD via targeted mutagenesis.
Supplementary information: The online version contains supplementary material available at 10.1007/s11032-025-01553-5.
期刊介绍:
Molecular Breeding is an international journal publishing papers on applications of plant molecular biology, i.e., research most likely leading to practical applications. The practical applications might relate to the Developing as well as the industrialised World and have demonstrable benefits for the seed industry, farmers, processing industry, the environment and the consumer.
All papers published should contribute to the understanding and progress of modern plant breeding, encompassing the scientific disciplines of molecular biology, biochemistry, genetics, physiology, pathology, plant breeding, and ecology among others.
Molecular Breeding welcomes the following categories of papers: full papers, short communications, papers describing novel methods and review papers. All submission will be subject to peer review ensuring the highest possible scientific quality standards.
Molecular Breeding core areas:
Molecular Breeding will consider manuscripts describing contemporary methods of molecular genetics and genomic analysis, structural and functional genomics in crops, proteomics and metabolic profiling, abiotic stress and field evaluation of transgenic crops containing particular traits. Manuscripts on marker assisted breeding are also of major interest, in particular novel approaches and new results of marker assisted breeding, QTL cloning, integration of conventional and marker assisted breeding, and QTL studies in crop plants.